Project 1: Targeting glypican-3 in hepatocellular carcinoma Heparan sulfate proteoglycans (HSPGs) are important modulators of signal transduction pathways during development and disease. They are cell-surface proteins that are modified by the addition of one or several glycosaminoglycan chains. Several HSPGs have been suggested as candidate targets for cancer therapy because of their high expression in certain tumor types. We have generated human and humanized antibodies targeting glypican-3 (GPC3) in hepatocellular carcinoma (HCC). We generated mouse monoclonal antibodies (e.g. YP7) that recognize a C-terminal site (511-560) in GPC3 [Phung et al., MAbs, PMID 22820551, 2012] . Furthermore, we generated two human monoclonal antibodies (HN3 and HS20). HN3 is a human heavy-chain antibody that recognizes a unique conformational epitope in the core protein of GPC3 and inhibits proliferation of HCC cells. The underlying mechanism of HN3 action involves inhibition of Yap signaling in liver cancer cells. HS20 preferentially recognizes the heparan sulfate chains of GPC3. The human antibody disrupts the interaction of Wnt3a and GPC3 and inhibits Wnt/beta-catenin signaling. The new antibodies exhibit significant inhibition of HCC xenograft tumor growth in mice and show potential for use as therapeutic candidates. In FY14, we summarized our work on the development of human monoclonal antibodies targeting GPC3 for liver cancer therapy and published two research reports about the HN3 and HS20 human antibodies [Feng et al., PNAS, PMID 23471984, 2013; Gao et al., Hepatology, PMID 24492943, 2014] and one invited review article about GPC3 therapeutic antibody development [Feng and Ho, FEBS Letters, PMID 24140348, 2014]. Project 2: Targeting mesothelin in mesothelioma, ovarian cancer and cholangiocarcinoma Mesothelin is expressed in mesothelioma, ovarian cancer, pancreatic cancer, lung cancer, gastric cancer, colorectal cancer, breast cancer and cholangiocarcinoma. The molecular interaction between mesothelin and MUC16 (also known as CA125) may facilitate the implantation and spread of tumors. We identified the functional binding domain (named IAB, 296-359) in mesothelin for MUC16 [Kaneko et al., JBC, PMID 19075018, 2009]. We generated two human monoclonal antibodies specific for mesothelin. The HN1 human antibody disrupts the mesothelin-MUC16 interaction and elicits antibody-dependent cell-mediated cytotoxicity (ADCC) against tumor cells. SD1 is a human heavy-chain antibody that recognizes a unique site (539-588) in mesothelin close to the cell surface and exhibits complement-dependent cytotoxicity (CDC) as well as ADCC against tumor cells. The new human antibodies show potential for use as cancer therapeutic candidates. In FY14, we summarized the SD1 antibody study and published a research article [Tang et al., Molecular Cancer Therapeutics, PMID 23371858, 2013]. In addition, we have also developed an immunocytokine based on IL12 and the SS1 Fv in collaboration with Ira Pastan (NCI). The new immunocytokine inhibits human mesothelioma grown in mice and provides an attractive addition to mesothelin-targeted cancer therapies. In FY14, we published our report about the immunocytokine targeting mesothelin-positive tumors [Kim et al., PLoS One, PMID 24260587, 2013].